title: Development and standardization of a monoclonal antibody-based rapid flow-through immunoassay for the detection of Aphanomyces invadans in the field.
abstract: A monoclonal antibody-based flow-through immunoassay (FTA) was developed using a nitrocellulose membrane placed on the top of adsorbent pads enclosed in a plastic cassette with a test zone at the center. The FTA could be completed within 10 min. Clear purple dots against a white background indicated the presence of Aphanomyces (A.) invadans. The FTA limit of detection was 7 μg/mL for A. invadans compared to 56 μg/mL for the immunodot. FTA and polymerase chain reaction (PCR) could detect A. invadans in fish tissue homogenates at a 10(-11) dilution compared to a 10(-8) dilution by immunodot. In fish suffering from natural cases of epizootic ulcerative syndrome (EUS) collected from Mangalore, India, FTA and PCR could detect A. invadans in 100% of the samples compared to 89.04% detected by immunodot. FTA reagents were stable and produced expected results for 4 months when stored at 4~8°C. This rapid test could serve as simple and cost-effective on-site screening tool to detect A. invadans in fish from EUS outbreak areas and in ports during the shipment of live or frozen fish.
authors.affiliation: Aquatic Health Management Laboratory, Department of Aquaculture, College of Fisheries, Karnataka Veterinary, Animal and Fisheries Sciences University, Mangalore 575002, India.
authors.forename: B
authors.initials: B
authors.lastname: Adil
authors.forename: K M
authors.initials: KM
authors.lastname: Shankar
authors.forename: B T Naveen
authors.initials: BT
authors.lastname: Kumar
authors.forename: Rajreddy
authors.initials: R
authors.lastname: Patil
authors.forename: Abhiman
authors.initials: A
authors.lastname: Ballyaya
authors.forename: K S
authors.initials: KS
authors.lastname: Ramesh
authors.forename: Sathish Rama
authors.initials: SR
authors.lastname: Poojary
authors.forename: Omkar V
authors.initials: OV
authors.lastname: Byadgi
authors.forename: Prabhugouda
authors.initials: P
authors.lastname: Siriyappagouder
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000378:metabolism; D044744:Aphanomyces / Q000302:isolation & purification*; D005393:Fish Diseases / Q000175:diagnosis* / Q000469:parasitology; D005399:Fishes; D007118:Immunoassay / Q000379:methods* / Q000662:veterinary; D007194:India; D007239:Infections / Q000175:diagnosis / Q000469:parasitology / Q000662:veterinary*; D016133:Polymerase Chain Reaction / Q000379:methods / Q000662:veterinary; D011933:Reagent Kits, Diagnostic / Q000592:standards* / Q000662:veterinary
chemical_list: D000911:Antibodies, Monoclonal; D011933:Reagent Kits, Diagnostic
keywords: diagnosis; epizootic ulcerative syndrome; fish; flow-through; monoclonal antibody